J. Electrochem. Sci. Eng. 14(1) (2024) 93-105; http://dx.doi.org/10.5599/jese.

2038

Open Access : : ISSN 1847-9286

www.jESE-online.org
Original scientific paper
Facile preparation of a sensitive electrochemical sensor with
good performance for determination of methionine
Peyman Mohammadzadeh Jahani1 and Somayeh Tajik2,
1School of Medicine, Bam University of Medical Sciences, Bam, Iran
2Research Centre for Tropical and Infectious Diseases, Kerman University of Medical Sciences,
Kerman, Iran
Corresponding author: tajik_s1365@yahoo.com; Tel.: +98-3431325700; Fax: +98-3431325700
Received: August 23, 2023; Accepted: November 10, 2023; Published: November 29, 2023

Abstract
In this work, a novel voltammetric sensor for the detection of methionine was designed and
prepared by using a carbon paste electrode (CPE) modified with ZnO hollow quasi-spheres
(ZnO hollow QSs) and 1-butyl-3-methylimidazolium hexafluorophosphate (BMIM.PF6). The
results by cyclic voltammetry showed that the prepared electrode (ZnO-BMIM.PF6/CPE)
effectively increased the oxidation peak current and reduced the oxidation peak potential of
methionine and had a suitable electrocatalytic activity for the oxidation of methionine.
Notably, the ZnO-BMIM.PF6/CPE exhibited high detection capability towards the
quantification of methionine in 0.1 M PBS (pH 7.0) over a concentration range from 0.04 to
330.0 µM with a limit of detection of 0.02 µM. More importantly, the effectiveness of the
ZnO-BMIM.PF6/CPE sensor was also confirmed in real samples (urine detection with
acceptable recoveries (98.0 to 102.7 %) and relative standard deviation values ≤ 3.3 %.
Keywords
Voltammetric sensor; carbon paste electrode; ZnO hollow quasi-spheres; 1-butyl-3-me-
thylimidazolium hexafluorophosphate

Introduction
Methionine is classified as a sulfur-containing amino acid because it contains a sulfur atom in this
chemical structure. Methionine is a primary source of sulfur in the diet, playing a vital role in
maintaining the health and integrity of various tissues, including the hair, skin, and nails [1]. Also,
methionine plays a crucial role in various biological processes, including protein synthesis, synthesis
of amino acids, such as cysteine, taurine, homocysteine, and glycine, transmethylation reaction, and
other physiological processes [2]. By increasing lecithin production in the liver, methionine can
indirectly reduce cholesterol levels [3]. In addition, methionine acts as a chelator for heavy metals
and functions as a powerful antioxidant for free radicals scavenging [3]. Methionine deficiency has
been studied in relation to various diseases, including toxaemia, Parkinson’s disease, and acquired

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immune deficiency syndrome (AIDS) [4]. In addition to this, methionine deficiencies can lead to hair
loss, weight loss, liver deterioration, impaired growth, depression, and muscle paralysis [5].
Therefore, developing an accurate and reliable analytical method for detecting methionine is crucial
due to its clinical and physiological significance. At present, several methods, such as capillary
electrophoresis [6], chromatography [7], colorimetry [8], fluorescence [9], chemiluminescence [10],
and so on, have been extensively used for the analysis of methionine. Although some of these
methods can be reliable, it is important to consider that they may require expensive and
sophisticated equipment as well as time-consuming procedures.
Electrochemical methods are still widely used and popular due to their distinct characteristics,
including fast response, low cost, versatility, simple operation, ease of miniaturization, and so on
[11-19]. Modified electrodes play a crucial role in enhancing the performance, sensitivity, and
selectivity of electrochemical sensors, allowing for more accurate and reliable detection of target
analytes [20-26]. Nanotechnology is a closely related field that deals with the study and
manipulation of materials and phenomena at the nanometer scale to create new materials,
structures, and functionalities. Nanotechnology opened up new possibilities for innovation in
various fields, including electronics, medicine, energy, materials science and more [27-36]. The
application of nanostructures for the modification of electrodes has gained significant attention in
recent years [37-41]. Nanostructured materials can offer enhanced properties such as high specific
surface area and high conductivity, making them ideal candidates for electrode modifications in
sensing applications. By providing higher sensitivity and selectivity, nanostructures improve the
performance of electrochemical sensors in detecting and measuring different species [42-44].
ZnO is regarded as a versatile material that has been extensively studied in a wide range of
applications in various fields, including catalysis [45], gas sensors [46], energy storage [47], electro-
chemical sensors and biosensors [48], water treatment [49], biomedicine [50], and etc. However,
researchers continuously explore innovative ways to synthesize ZnO nanostructures with desired
properties to unlock their potential fully. By manipulating the synthesis process, researchers can
control the shape, size, and morphology of ZnO nanostructures, leading to significant changes in
their physical and chemical characteristics. This control over nanostructure design opens avenues
for tailoring ZnO properties to meet specific application requirements. In recent years, ZnO hollow
nanostructures have gained significant attention in scientific research and technological applica-
tions [51-53]. The unique properties of ZnO hollow nanostructures, including low density, porous
structure, and high specific surface area, make them promising candidates for the development of
high-performance electrochemical sensors.
Ionic liquids (ILs) are non-molecular ionic compounds composed of oppositely charged ions
arranged in a crystal lattice structure, and they exhibit distinct properties different from molecular
compounds. The diverse combinations of cations and anions allow for the creation of ILs with
tailored properties and functionalities [54]. ILs have gained significant attention in various fields,
notably electrochemistry, because of their thermal and chemical stability, high conductivity, wide
potential window, and low vapor pressure [55]. The combination of nanomaterials with ILs has
shown great potential in the fabrication of electrochemical sensors. By creating the synergistic
effects of nanomaterials and ILs, researchers can design and fabricate innovative electrochemical
sensors with improved performance, sensitivity, and selectivity. This opens up new possibilities for
applications in fields such as environmental monitoring, healthcare diagnostics, and industrial
process control [56,57].

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Herein, we developed a high-performance modified CPE based on ZnO hollow QSs-BMIM.PF6 for
detection of methionine. The ZnO-BMIM.PF6 modified CPE reduces the overpotential and enhances
the oxidation peak current for the effective electrochemical detection of methionine. Furthermore,
the modified CPE provided acceptable results for the detection of methionine in real samples.

Experimental
Instruments and materials
All electrochemical studies and measurements were done using a potentiostat/galvanostat
device (Metrohm Autolab – PGSTAT302N (Utrecht, The Netherlands)), controlled by the GPES
4.9004 software. The electrochemical tests were performed in a typical three-electrode setup by
using reference electrode (RE) (Ag/AgCl/KCl (3 M)), counter electrode (CE) (platinum), and working
electrode (modified CPE). All solvents and chemicals were commercially available (Merck and Sigma-
Aldrich companies) with analytical grade and used directly without further purification.
The synthesis and characterization of ZnO hollow QSs were reported in our previous work [58].
Figure 1 shows the FE-SEM image of ZnO hollow QSs.

Figure 1. FE-SEM image of ZnO hollow QSs

Preparation of ZnO-BMIM.PF6/CPE
The ZnO-BMIM.PF6 modified CPE with a mass of 0.5 g was achieved by hand-mixing 0.48 g of
graphite powder and 0.02 g of ZnO hollow QSs for 5 min until a homogeneous blend was formed.
Then, paraffin oil and BMIM.PF6 in the ratio 3:1 was added to the blend of graphite and ZnO hollow
QSs, which was mixed again for at least 30 min to obtain the ZnO-BMIM.PF6 modified carbon paste.
Finally, the modified paste was packed into the glass tube cavity. The electrical contact was

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established through a conductive copper wire. Also, the surface of the prepared electrode
(ZnO-BMIM.PF6/CPE) was polished on a smooth paper to obtain a shiny and smooth appearance.
To calculate the electrochemically active surface area (EASA) of the unmodified CPE and
ZnO-BMIM.PF6/CPE, the CVs were recorded at different scan rates in 0.1 M KCl solution containing
1.0 mM K3[Fe(CN)6] as a redox probe. Using the Randles–Ševčik equation, the value of the ESCA for
ZnO-BMIM.PF6/CPE (0.297 cm2) was found 3.3 times greater than unmodified CPE.

Results and discussion

Electrocatalytic response of ZnO-BMIM.PF6/CPE towards methionine
The effect of pH values (from 2.0 to 9.0) of the supporting electrolyte (0.1 M PBS) on methionine's
electrochemical oxidation was studied using the ZnO-BMIM.PF6 modified CPE via DPV technique. It was
observed that by changing the pH value of PBS, the prepared electrode showed different voltam-
mograms for oxidation of methionine. The peak potential and peak current from the oxidation of
methionine showed a strong dependence on pH. By increasing the pH from lower to higher values, the
anodic peak potential of methionine was shifted towards the negative potentials. Also, the Ipa of
methionine gradually increased with the increase of pH from 2.0 to 7.0 and then decreased. The
maximum Ipa was obtained at pH 7.0. Therefore, pH 7.0 was used for further electrochemical studies.
To assess the electrocatalytic activity of the IL (BMIM.PF6) and as-prepared ZnO, the electro-
chemical responses of methionine on unmodified CPE and modified CPE were examined by cyclic
voltammetry (CV). Figure 2 shows the cyclic voltammograms from the response of unmodified CPE
(voltammogram a) and ZnO-BMIM.PF6/CPE (voltammogram b) towards the 150.0 µM methionine in
0.1 M PBS (pH 7.0).

E / mV vs. Ag/AgCl/KCl
Figure 2. CVs of unmodified CPE (a) and ZnO-BMIM.PF6/CPE (b) in 0.1 M PBS (pH 7.0) containing 150.0 µM
methionine at a scan rate of 50 mV s-1

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As can be seen, a broad oxidation peak with a low anodic peak current (Ipa) was shown for
unmodified CPE. The ZnO-BMIM.PF6/CPE clearly improves the oxidation of methionine, as evident
from the increase of the Ipa (from 3.5 to 13.0 µA) and decrease of the overpotential (from 950 to
850 mV) when compared with unmodified CPE. This result could be related to the electrocatalytic
effect of the IL and ZnO NPs. In addition, the absence of any reduction peak on the reverse scan
revealed the irreversible oxidation of methionine over unmodified and modified CPE.
Effect of scan rate on the oxidation reaction of methionine
To investigate the effect of scan rate, CVs of the ZnO-BMIM.PF6/CPE were recorded at different
scan rates (10 to 250 mV/s) in 0.1 M PBS containing 100.0 µM methionine (Figure 3). An increase in
the anodic peak current (Ipa) with an increase in scan rate can be observed. Also, from the obtained
voltammograms, it was possible to observe a linear dependence between Ipa of methionine and the
square root of scan rate (1/2) (Ipa = 1.89011/2 -2.9739) (Figure 3 Inset). This observation suggests
that the oxidation reaction is controlled by the diffusion of methionine species from the bulk
solution to the surface of ZnO-BMIM.PF6/CPE.

E / mV vs. Ag/AgCl/KCl
Figure 3. CVs of ZnO-BMIM.PF6/CPE performed at different scan rates (from a: 10 to e: 250 mV s-1) in
0.1 M PBS (pH 7.0) containing 100.0 µM methionine. Inset: the linear dependence between Ipa vs. 1/2

Chronoamperometric measurements of methionine at ZnO-BMIM.PF6/CPE

To measure the diffusion coefficient (D) of methionine, the chronoamperometric responses of
ZnO-BMIM.PF6/CPE were plotted for different concentrations of methionine from 0.1 to 1.7 mM at the

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fixed potential of 0.9 V (Figure 4). The current-time (I-t) curves reflect the change in concentration
gradient of the electroactive species (methionine) in the vicinity of the electrode surface as time
progresses. To determine the D, the Cottrell curves (I versus t-1/2) were plotted over a certain range of
time for different concentrations of methionine (Figure 4A). Then, the slope of the obtained Cottrell
curves was plotted vs the different concentrations of methionine (Figure 4B) and a straight line with a
slope of 19.7 µA s1/2 mM-1 was obtained. From the slope of the resulting plot and using Cottrell's
equation, the D of methionine on the surface of ZnO-BMIM.PF6/CPE was found to be 1.6×10-5 cm2 s-1.

Figure 4. Chronoamperometric responses of ZnO-BMIM.PF6/CPE in 0.1 M PBS (pH 7.0) containing

different concentrations of methionine from a: 0.1 to 1.7 mM. Inset A: the linear dependence between
Ipa / µA vs. t-1/2 / s-1/2) and Inset B: linear dependence between slope values of I-t-1/2 plots vs. methionine
concentrations

Quantitative analysis of methionine by DPV

To study the detection efficiency of ZnO-BMIM.PF6/CPE, the DPV measurements were performed
with the successive addition of methionine (0.04 to 330.0 µM) in 0.1 M PBS (pH 7.0) in the following
conditions: step potential 0.01 V and pulse amplitude 0.025 V (Figure 5). From the recorded
voltammograms, the increase of the Ipa is proportional to the increase of methionine concentration in
a wide range from 0.04 to 330.0 µM. Furthermore, the linear dependence between the enhanced Ipa
of methionine and its concentration is presented in the Inset of Figure 5. This dependence can be
expressed by I = 0.0812CMethionine + 0.8557 with a correlation coefficient of 0.999. The LOD was
calculated according to the ensuing formula 3Sb/m, where Sb denotes the standard deviation of the

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blank (PBS) signal (obtained based on 12 measurements on the blank solution), and m denotes the
slope of the corresponding calibration curve, and it was found to be 0.02 µM. The limit of quanti-
fication was found to be 0.04 µM. Table 1 lists the comparative characteristics of the as-prepared
sensor with those of previously reported sensors for the determination of methionine.

E / mV vs. Ag/AgCl/KCl
Figure 5. DPVs of ZnO-BMIM.PF6/CPE performed in 0.1 M PBS (pH 7.0) containing different concentrations
of methionine (from a: 0.04 to m: 330.0 µM). Inset: the linear dependence between Ipa vs. methionine
concentration

Table 1. Comparative results of ZnO-BMIM.PF6/CPE based methionine sensor with previously reported sensors
Electrochemical sensor Linear range, µM LOD, µM Sensitivity Ref.
Pt-doped TiO2 nanoparticles )NPs)
carbon nanotubes (CNTs)/glassy 0.5 - 100 0.1 29.085 µA µM-1 cm-2 [1]
carbon electrode (GCE)
Colloidal gold-cysteamine/CPE 1.0 - 100 0.59 - [12]
Fullerene-C60/Au electrode - 8.2 50 mA M-1 [59]
Ni-doped carbon ceramic electrode 2 - 90 2 5.6 nA μM-1 [60]
Graphitic carbon nitride
0.1 - 200 0.32×10-3 1.16 µA µM-1 cm-2 [61]
nanosheets/GCE
ZnO-BMIM.PF6/CPE 0.04 - 330.0 0.02 0.0812 µA µM-1 This work

Stability and reproducibility studies of ZnO-BMIM.PF6/CPE sensor towards the determination of

methionine
Studies related to the stability of ZnO-BMIM.PF6/CPE sensors were performed by recording the
current response of the designed sensor towards 75.0 µM methionine over 20 days. The results

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showed that the electrode response retained 95.9 % of its initial value after 20 days. These results
indicated that the designed sensor has good stability.
Also, the reproducibility of the prepared sensor (ZnO-BMIM.PF6/CPE) was evaluated by recording
the current response of five electrodes prepared independently under the same conditions. All five
prepared electrodes showed almost the same responses and the relative standard deviation (RSD)
was 2.7 % in the determination of 75.0 µM methionine.
Interferences studies
The effect of the possible interferences from some species such as Na+, Ca2+, Mg2+, NH4+, Al3+, Cl-,
SO42-, S2-, glucose, acetaminophen, epinephrine, norepinephrine, uric acid, tryptophan, glycine,
phenylalanine, and L-serine on the electrochemical response of methionine was evaluated at the
surface of ZnO-BMIM.PF6/CPE sensor. It was observed that these species did not show significant
interference for the determination of methionine (no signal change more than ± 5 %). These results
confirmed that the developed sensor has good selectivity for the determination of methionine.
Methionine analysis in real samples
To evaluate the practical performance of the developed sensor (ZnO-BMIM.PF6/CPE), the deter-
mination of methionine in the urine sample was conducted. The standard addition method was
employed for the analysis of methionine by the DPV technique. By adding the known concentrations
of methionine to the urine sample, measurements were performed. The recovery and RSD values
are summarized in Table 2. The summarized results in Table 1 show acceptable recovery values
(between 98.0 and 102.7 %) and RSD values (n = 5) of ≤3.3 %, which confirm that the developed
sensor could be used for real-time analysis.

Table 2. Real sample analysis for the determination of methionine spiked into the urine samples
Amount, µM
Sample Recovery, % RSD, %
Spiked Found
0 - - -
5.0 4.9±0.05 98.0 3.3
Urine 7.5 7.7±0.04 102.7 2.9
10.0 10.1±0.06 101.0 1.7
12.5 12.4±0.05 99.2 2.4

Conclusions
In this study, the efficient and accurate detection of methionine was reported based on ZnO hollow
QSs-BMIM.PF6 modified CPE. The obtained results demonstrated that the ZnO-BMIM.PF6/CPE sensor
was well developed and showed an enhanced electrochemical response towards methionine
oxidation. The ZnO-BMIM.PF6/CPE can be used to determine methionine in the concentration from
0.04 to 330.0 M with an LOD of 0.02 µM. Finally, excellent precision (RSD ≤3.3 %) and accuracy
(recovery for spiked samples ranging from 98.0 to 102.7 %) were obtained.

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